Application of r-ketamine and salt thereof as pharmaceuticals

ABSTRACT

Provided is a novel compound having rapid and long-lasting therapeutic effects on diseases exhibiting depressive symptoms. Specifically, provided are an agent for prevention and/or treatment of a depressive symptom, consisting of R(−)-ketamine or a pharmacologically acceptable salt thereof, and a pharmaceutical composition for prevention and/or treatment of a depressive symptom, comprising R(−)-ketamine or a pharmacologically acceptable salt thereof in an effective amount for reducing a depressive symptom, and being substantially free of S(+)-ketamine, and a pharmacologically acceptable salt thereof.

TECHNICAL FIELD

The present invention relates to a pharmaceutical for prevention and/ortreatment of psychiatric diseases, preferably diseases exhibitingdepressive symptoms. More specifically, the present invention relates toan antidepressant including R-ketamine or a pharmaceutically acceptablesalt thereof, and to a pharmaceutical composition for prevention and/ortreatment of diseases exhibiting depressive symptoms, includingR-ketamine or a pharmaceutically acceptable salt thereof, and beingsubstantially free of S-ketamine or a pharmaceutically acceptable saltthereof.

BACKGROUND ART

Along with changes in social life style and aging of society, variousdiseases such as psychiatric diseases and neurological diseases tend toincrease as a whole. For example, high incidences of depression andschizophrenia, which are major psychiatric diseases, have become aserious problem from the viewpoint of medical economy as well. Inaddition, obsessive-compulsive disorder is an anxiety disorder involvingobsessions and compulsions. In treatment of the psychiatric diseasessuch as depression, schizophrenia, anxiety disorders, and autismspectrum disorder, medication is essential, and an antidepressant (e.g.,a tricyclic antidepressant, a selective serotonin reuptake inhibitor,and a serotonin and norepinephrine reuptake inhibitor), an antipsychotic(e.g., a phenothiazine-based compound, a butyrophenone-based compound, abenzamide-based compound, an iminodibenzyl compound, a thiepin-basedcompound, an indole-based compound, and a serotonin/dopamine receptorantagonist), and an anti-anxiety drug are administered. However, thosedrugs used actually in a clinical field are effective for some patientsand some symptoms, but patients for whom the drugs are ineffective,so-called treatment-resistant patients are also known to exist. Thus,there is a strong demand for development of a novel therapeutic drug. Itis hard to say that the existing drugs exhibit sufficient therapeuticeffects on those psychiatric diseases. In reality, there aresubstantially no effective prevention and treatment methods at present.

One of the major problems in treatment of depression is that there arelimitations on effects of the antidepressant and effects of its adjuvanttherapy. It takes several weeks or more for the current antidepressantsto express their drug efficacy. In addition, there existtreatment-resistant patients for whom those antidepressants areineffective. Therefore, it is also said that only 50% of patients withdepression reach remission. In addition, when a dose of theantidepressant is increased for achieving remission, a patient suffersfrom various side effects accordingly. Further, depression is one of thecause of suicide. Depression in elderly peoples is known to increase therisk if incident dementia, in particular of Alzheimer's disease andvascular dementia (Non Patent Literature 1).

In recent research, growing evidence suggests that abnormality inglutamatergic transmission, in particular, glutamatergicneurotransmission via an N-methyl-D-aspartate (hereinafter abbreviatedas NMDA) receptor is associated with pathophysiology of mood disorderssuch as major depressive disorder (hereinafter abbreviated as MDD) andbipolar disorder. The MNDA receptor also plays key roles in neurobiologyand treatment of MDD as well (Non Patent Literature 2).

It has been reported that an NMDA receptor antagonist ketamine exhibitsrapid and robust antidepressant effects on treatment-resistant patientswith MDD and depressive symptoms of treatment-resistant bipolar disorder(Non Patent Literatures 3 to 5). In addition, it has been reported thatketamine is also effective for treatment-resistant obsessive-compulsivedisorder and treatment-resistant posttraumatic stress disorder(hereinafter abbreviated as PTSD) (Non Patent Literatures 6 to 8).Ketamine has also been reported to have an effect of inhibiting suicideideation (Non Patent Literature 9). Further, ketamine treatment in anadult with autism spectrum has been reported (Non Patent Literature 10).Ketamine, which was a compound developed as an anesthetic in 1962,started to be applied clinically in 1965. However, ketamine isdesignated as a controlled substance because of its problems ofpsychotic symptoms such as hallucination and delusion, and drugdependence. At present, ketamine is used as an anesthetic and fortreatment of chronic pain in a clinical field.

It has been reported that clinical antidepressant effects of ketaminelast for a short period of from 1 to 2 days starting from several hoursafter its single administration. Meanwhile, it has been reported thatthe effects may last over 2 weeks or more (Non Patent Literatures 3, 4,and 11). In addition, it has been reported that ketamine haspsychotomimetic effects as side effects, and antidepressant effects ofketamine were not present until after the side effects had disappeared(Non Patent Literatures 3 and 4).

Ketamine (or sometimes referred to as RS(+/−)-ketamine) is a racemicmixture containing equal amounts of R(−)-ketamine and S(+)-ketamine.R(−)-ketamine and S(+)-ketamine are also called R-isomer and S-isomer ofketamine, respectively. S(+)-ketamine has approximately 4-fold greateraffinity for the NMDA receptor than R-isomer (Non Patent Literature 12).Further, S(+)-ketamine has an approximately 3-to 4-fold anestheticeffect as compared to R-isomer, and has greater psychotomimetic sideeffects than R-isomer (Non Patent Literature 12). As described above,the potency of psychotomimetic effects of ketamine is correlated withthe potency of blockade of the NMDA receptor (Non Patent Literature 12).A positron emission tomography (PET) study in healthy volunteersdemonstrated that psychotomimetic doses of S(+)-ketamine (i.e.,intravenous infusion of 15 mg for 5 min, then infusion of the dose(0.014 to 0.02 mg/kg/min for 53 min) increased cerebral metabolic ratesof glucose (hereinafter abbreviated as CMRglu) markedly in the frontalcortex and thalamus (Non Patent Literature 13). In contrast, equimolardoses of R(−)-ketamine tended to decrease CMRglu across brain regions,and did not produce psychotic symptoms, but a state of relaxation and afeeling of well being (Non Patent Literature 13).

As described above, it is generally understood that both analgesiceffects and psychotomimetic effects of ketamine are mediated primarilyvia the blockade of the NMDA receptor. S-isomer of ketamine has highaffinity for the NMDA receptor. Thus, it is considered that thoseeffects of ketamine are caused primarily by S-isomer.

At present, ketamine is one of the drugs that have attracted attentionfor treatment of treatment-resistant patients with MDD, depressivesymptoms of treatment-resistant bipolar disorder, treatment-resistantobsessive-compulsive disorder, and treatment-resistant PTSD (Non PatentLiteratures 5 to 12). A previous case report showed that antidepressanteffects of S(+)-ketamine (0.25 mg/kg, i.v.) in treatment-resistantpatients with MDD were weaker than those of RS(+/−)-ketamine (0.5 mg/kg,i.v.) (Non Patent Literature 14). Further, an open label study (NonPatent Literature 15) and a case report (Non Patent Literature 16)showed that effective oral doses of RS(+/−)-ketamine and S(+)-ketaminein patients with depression were 0.5 mg/kg and 1.25 mg/kg, respectively.In addition, intranasal administration of ketamine showed antidepressanteffect in treatment-resistant patients with MDD (Patent Literature 1 andNon Patent Literature 17).

CITATION LIST Patent Literature

[PTL 1] International Patent Publication No. WO 2007/111880 A2

[PTL 2] U.S. Pat. No. 6,040,479 (A)

Non Patent Literature

[NPL 1] Diniz BD, Butters MA, Albert SM, Dew MA and Reynolds CF (2013)Late-life depression and risk of vascular dementia and Alzheimer'sdisease: systematic review and meta-analysis of community-based cohortstudies. B. J. Psychiatry 202: 329-335.

[NPL 2] Hashimoto K (2009) Emerging role of glutamate in thepathophysiology of major depressive disorder. Brain Res. Rev. 61:105-23.

[NPL 3] Berman R M, Cappiello A, Anand A, Oren D A, Heninger G R,Charney D S, Krystal J H (2000) Antidepressant effects of ketamine indepressed patients. Biol. Psychiatry 47:351-4.

[NPL 4] Zarate C A, Jr, Singh J B, Carlson P J. Brutsche N E, Ameli R,Luckenbaugh D A, Charney D S, Manji H K (2006) A randomized trial of anN-methyl-D-aspartate antagonist in treatment-resistant major depression.Arch. Gen. Psychiatry 63:856-64.

[NPL 5] Diazgranados N, Ibrahim L. Brutsche NE. Newberg A, Kronstein P,Khalifc S, Kammerer W A, Quezado Z, Luckenbaugh D A, Salvadore G,Machado-Vieira R, Manji H K, Zarate C A Jr. (2010) A randomized add-ontrial of an N-methyl-D-aspartate antagonist in treatment-resistantbipolar depression. Arch. Gen. Psychiatry 67:793-802.

[NPL 6]Bloch M H, Wasylink S, Landeros-Weisenberger A, Panza K E,Billingslea E, Leckman J F, Krystal J H, Bhagwagar Z, Sanacora G,Pittenger C (2012) Effects of ketamine in treatment-refractoryobsessive-compulsive disorder. Biol. Psychiatry 72(11):964-970.

[NPL 7] Rodriguez C I, Kegeles L S, LevinsonA, Feng T, Marcus S M,Vermes D, Flood P, Simpson H B (2013) Randomized Controlled CrossoverTrial of Ketamine in Obsessive-Compulsive Disorder: Proof-of-Concept.Neuropsychopharmacology 38:2475-83.

[NPL 8] Feder A, Parides M K, Murrough J W, Perez A M, Morgan J E,Saxena S. Kirkwood K, Aan Het Rot M, Lapidus K A, Wan L B, Iosifescu D,Charney D S (2014) Efficacy of intravenous ketamine for treatment ofchronic posttraumatic stress disorder: a randomized clinical trial. JAMAPsychiatry 71:681-688.

[NPL 9] DiazGranados N, Ibrahim L A, Brutsche N E, Ameli R, Henter I D,Luckenbaugh D A, Machado-Vicira R. Zarate C A Jr (2010) Rapid resolutionof suicidal ideation after a single infusion of an N-methyl-D-aspartateantagonist in patients with treatment-resistant major depressivedisorder. J Clin. Psychiatry 71(12):1605-11.

[NPL 10] Wink LK1, O′Melia A M, Shaffer R C, Pedapati E, Friedmann K,Schaefer T, Erickson C A (2014) Intranasal ketamine treatment in anadult with autism spectrum disorder. J Clin. Psychiatry 75(8):835-6.doi: 10.4088/JCP.13cr08917.

[NPL 11] Krystal J H, Sanacora G, Duman R S (2013) Rapid-actingglutamatergic an-tidepressants: the path to ketamine and beyond. Biol.Psychiatry 73:1133-41.

[NPL 12] Domino EF (2010) Taming the ketamine tiger. 1965.Anesthesiology 113:678-86.

[NPL 13] Vollenweider F X. Leenders K L, OEye I, Hell D, Angst J (1997)Differential psychopathology and patterns of cerebral glucoseutilization produced by (S)- and (R)-ketamine in healthy volunteersusing positron emission tomography (PET). Eur. Neuropsychopharmacol. 7:25-38.

[NPL 14] Paul R, Schaaff N, Padberg F, Moeller H J, Frodl T (2009)Comparison of racemic ketamine and S-ketamine in treatment-resistantmajor depression: report from two cases. World J. Biol.Psychiatry 10:241-244.

[NPL 15] Paslakis G, Gilles M, Meyer-Lindenberg A, Deuschle M (2010)Oral administration of the NMDA receptor antagonist S-ketamine as add-ontherapy of depression: a case series. Pharmacopsychiatry 43: 33-35.

[NPL 16] Irwin S A, Iglewicz A, Nelesen R A,Lo JY. Carr C H, Romero S D,Lloyd L S (2013) Daily oral ketamine for the treatment of depression andanxiety in patients receiving hospice care: A 28-day open-labelproof-of-concept trial. J. Palliat. Med. 16: 958-965.

[NPL 17] Lapidus K A, Levitch C F, Perez A M, Brallier J W, Parides M K,Soleimani L, Feder A, Iosifescu D V, Charney D S, Murrough J W (2014) Arandomized controlled trial of intranasal ketamine in major depressivedisorder. Biol. Psychiatry 2014 Apr 3. pii: S0006-3223(14)00227-3.doi:10.1016/j.biopsych.2014.03.026. [Epub ahead of print]

[NPL 18] Li S X, Fujita Y, Zhang JC, Ren Q, Ishima T, Wu J, Hashimoto K(2014) Role of the NMDA receptor in cognitive deficits, anxiety anddepressive-like behavior in juvenile and adult mice after neonataldexamethasone exposure. Neurobiol. Dis. 62: 124-134.

[NPL 19] Golden S A, Covington H E, III, Berton O, Russo S J (2011) Astandardized protocol for repeated social defeat stress in mice. Nat.Protoc. 6: 1183-1191.

SUMMARY OF INVENTION Technical Problem

It has been reported that the NMDA receptor antagonist ketamine exhibitsrapid antidepressant effects in treatment-resistant patients withdepression. The glutamatergic neurotransmission via the NMDA receptor isconsidered to be involved in depression, ketamine includes opticalisomers, i.e., S-isomer and R-isomer, and S-isomer has higher affinityfor the NMDA receptor than R-isomer. Thus, S-isomer or a racemic mixturehas been used for research on treatment of depression with ketamine.However, ketamine has problems of side effects including psychoticsymptoms such as hallucination and delusion, and dependence, and isdesignated as a controlled substance. Accordingly, it is difficult topractically use ketamine in a clinical field.

An object of the present invention is to provide a novel compound havingrapid and long-lasting antidepressant effects on diseases exhibitingdepressive symptoms, such as depression, bipolar disorder,obsessive-compulsive disorder, PTSD, and autism spectrum disorder.

Solution to Problem

The inventors of the present invention have made intensive studies inorder to achieve the above-mentioned object. In the studies. theinventors have focused attention on R(−)-ketamine, which has not beenused for research on antidepressant effects of ketamine heretofore. Inaddition, in research using a mouse model of depression, the inventorshave found that R(−)-ketamine exhibits more potent antidepressanteffects on the depression-like symptoms of the mouse model at a juvenilestage than S(+)-ketamine, and the effects last for a longer period. Theinventors also found in social defeat stress model mice thatR(−)-ketamine showed more potent and long lasting anti-depressant effectcompared to S(+)-ketamine. Furthermore, administration of S(+)-ketamineinduced some side effects such as a hyperlocomotion, prepulse inhibitiondeficit, and drug dependence, while administration of R(−)-ketamine didnot. Since R-isomer of ketamine has low affinity for an NMDA receptor ascompared to its S-isomer. the R-isomer is considered to have lesspsychotomimetic effects as side effects and to hardly produce drugdependence. The present invention has been accomplished based on thosefindings.

That is, the present invention relates to an agent for prevention and/ortreatment of a depressive symptom, consisting of R(−)-ketamine or apharmacologically acceptable salt thereof.

The present invention also relates to the agent, in which the depressivesymptom is a depressive symptom in depression in children or adults.

The present invention also relates to a pharmaceutical composition forprevention and/or treatment of a depressive symptom, comprisingR(−)-ketamine or a pharmacologically acceptable salt thereof in aneffective amount for reducing a depressive symptom, and beingsubstantially free of S(+)-ketamine or a pharmacologically acceptablesalt thereof.

The present invention also relates to the pharmaceutical composition, inwhich the depressive symptom is a depressive symptom in depression inchildren or adults.

The present invention also relates to a pharmaceutical composition forprevention and/or treatment of obsessive-compulsive disorder, comprisingR(−)-ketamine or a pharmacologically acceptable salt thereof in aneffective amount for reducing a depressive symptom inobsessive-compulsive disorder, and being substantially free ofS(+)-ketamine or a pharmacologically acceptable salt thereof.

The present invention also relates to a pharmaceutical composition forprevention and/or treatment of posttraumatic stress disorder, comprisingR(−)-ketamine or a pharmacologically acceptable salt thereof in aneffective amount for reducing a depressive symptom in posttraumaticstress disorder, and being substantially free of S(+)-ketamine or apharmacologically acceptable salt thereof.

The present invention also relates to a method of treating a depressivesymptom, comprising administering R(−)-ketamine or a pharmacologicallyacceptable salt thereof to a subject, wherein the subject has beendiagnosed with having a depressive symptom, in an amount effective totreat the depressive symptom.

The present invention also relates to the method, in which the subjecthas been diagnosed with depression in children or adults

The present invention also relates to the method, in which the subjecthas been diagnosed with obsessive-compulsive disorder.

The present invention also relates to the method, in which the subjecthas been diagnosed with posttraumatic stress disorder.

The present invention also relates to a method of treating a depressivesymptom, comprising administering a pharmaceutical composition to asubject, wherein the pharmaceutical composition comprises R(−)-ketamineor a phaimaceutically acceptable salt thereof in an amount effective totreat a depressive symptom, and a pharmaceutically acceptable carrier,and being substantially free of S(+)-ketamine or a pharmacologicallyacceptable salt thereof, and wherein the subject has been diagnosed withhaving a depressive symptom.

The present invention also relates to the method, in which the subjecthas been diagnosed with depression in children or adults.

The present invention also relates to the method, in which the subjecthas been diagnosed with obsessive-compulsive disorder.

The present invention also relates to the method, in which the subjecthas been diagnosed with posttraumatic stress disorder.

The present invention also relates to use of R(−)-ketamine for themanufacture of a pharmaceutical composition for treating a depressivesymptom.

The present invention also relates to the use, in which the depressivesymptom accompanies depression in children or adults.

The present invention also relates to the use, in which the depressivesymptom accompanies obsessive-compulsive disorder.

The present invention also relates to the use, in which the depressivesymptoms accompanies posttraumatic stress disorder.

ADVANTAGEOUS EFFECTS OF INVENTION

R(−)-ketamine or a pharmaceutically acceptable salt thereof has rapidand long-lasting antidepressant effects and less side effects, and henceis effective for prevention and/or treatment of psychiatric diseasesexhibiting depressive symptoms. Accordingly, the agent consisting ofR(−)-ketamine or a pharmacologically acceptable salt thereof, and thepharmaceutical composition including R(−)-ketamine or apharmacologically acceptable salt thereof, and being substantially freeof S(+)-ketamine or a pharmacologically acceptable salt thereof areuseful as novel pharmaceuticals in the field of prevention and/ortreatment of psychiatric diseases exhibiting depressive symptoms.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is a diagram illustrating the preparation of R(−)- andS(+)-ketamine hydrochloride (Ketamine HCl) from RS(+/−)-ketamine usingD(−)- and L(+)-tartaric acid (Tartaric acid), respectively.

FIG. 2A is a diagram illustrating a test protocol for investigatingantide-pressant effects of R(−)- and S(+)-ketamine. Tests were performedusing mice treated neonatally with dexamethasone (hereinafter referredto as DEX-treated mice) as a new animal model of depression. In FIG. 2A,DEX means dexamethasone, LMT means a locomotion test. TST means a tailsuspension test. FST means a forced swimming test, and SPT means a 1%sucrose preference test. (Example 1)

FIG. 2B is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the LMTthe day after the injection of ketamine. In FIG. 2B, R-Ket and S-Ketrepresent DEX-treated mouse groups injected with R(−)-ketamine andS(+)-ketamine, respectively, Saline represents a DEX-treated mouse groupinjected with saline, and Cont represents a control mouse group injectedwith saline. The ordinate axis of FIG. 2B indicates locomotions(count/60 min). (Example 1)

FIG. 2C is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the TSTthe day (27 hours) after the injection of ketamine. In FIG. 2C, R-Ketand S-Ket represent DEX-treated mouse groups injected with R(−)-ketamineand S(+)-ketamine, respectively, Saline represents a DEX-treated mousegroup injected with saline, and Cont represents a control mouse groupinjected with saline. The ordinate axis of FIG. 2C indicates immobilitytimes (sec) in the TST. (Example 1)

FIG. 2D is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the FSTthe day (29hours) after the injection of ketamine. In FIG. 2D, R-Ket andS-Ket represent DEX-treated mouse groups injected with R(−)-ketamine andS(+)-ketamine, respectively, Saline represents a DEX-treated mouse groupinjected with saline, and Cont represents a control mouse group injectedwith saline. The ordinate axis of FIG. 2D indicates immobility times(sec) in the FST. (Example 1)

FIG. 2E is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the SPT2 days after the injection of ketamine. In FIG. 2E, R-Ket and S-Ketrepresent DEX-treated mouse groups injected with R(−)-ketamine andS(+)-ketamine, respectively. Saline represents a DEX-treated mouse groupinjected with saline, and Cont represents a control mouse group injectedwith saline. The ordinate axis of FIG. 2E indicates sucrose preferences(%) in the SPT. (Example 1)

FIG. 2F is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the TST7 days after the injection of ketamine. In FIG. 2F, R-Ket and S-Ketrepresent DEX-treated mouse groups injected with R(−)-ketamine andS(+)-ketamine, respectively. Saline represents a DEX-treated mouse groupinjected with saline, and Cont represents a control mouse group injectedwith saline. The ordinate axis of FIG. 2F indicates immobility times(sec) in the TST. (Example 1)

FIG. 2G is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the DEX-treated mice investigated by the FST7 days after the injection of ketamine. In FIG. 2G, R-Ket and S-Ketrepresent DEX-treated mouse groups injected with R(−)-ketamine andS(+)-ketamine, respectively. Saline represents a DEX-treated mouse groupinjected with saline, and Cont represents a control mouse group injectedwith saline. The ordinate axis of FIG. 2G indicates immobility times(sec) in the FST. (Example 1)

FIG. 3A is a diagram illustrating a test protocol for investigatingantidepressant effects of R(−)- and S(+)-ketamine in social defeatstress mice. The social defeat stress mice were prepared by bringingC57/B6 male mice into contact with ICR male mice for 10 consecutive days(D1 to 10). After that, any one of R(−)-ketamine and S(+)-ketamine wasinjected and various tests were performed on day 1, day2, day 6, and day7 (P1, P2, P6, and P7) after the injection. In FIG. 3A, R-Ket and S-Ketrepresent social defeat stress mouse groups injected with R(−)-ketamineand S(+)-ketamine, respectively. Saline represents a social defeatstress mouse group injected with saline. In FIG. 3A, LMT means alocomotion test, TST means a tail suspension test. FST means a forcedswimming test, and SPT means a 1% sucrose preference test. (Example 2)

FIG. 3B is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe SPT 1 day (P1) after the injection of ketamine. In FIG. 3B, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3B indicates sucrose preferences (%) in the SPT. (Example 2)

FIG. 3C is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe LMT 2 days(P2) after the injection of ketamine. In FIG. 3C, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3C indicates locomotions (count/60min) in the LMT. (Example 2)

FIG. 3D is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe TST 2 days (P2) after the injection of ketamine. In FIG. 3D, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3D indicates immobility times (sec) in the TST. (Example 2)

FIG. 3E is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe FST 2 days (P2) after the injection of ketamine. In FIG. 3E, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3E indicates immobility times (sec) in the FST. (Example 2)

FIG. 3F is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe SPT 6 days (P6) after the injection of ketamine. In FIG. 3F, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3F indicates sucrose preferences (%) in the SPT. (Example 2)

FIG. 3G is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe TST 7 days (P7) after the injection of ketamine. In FIG. 3G, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3G indicates immobility times (sec) in the TST. (Example 2)

FIG. 3H is a graph showing results of the antidepressant effects ofR(−)- and S(+)-ketamine in the social defeat stress mice investigated bythe FST 7 days (P7) after the injection of ketamine. In FIG. 3H, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. The ordinate axisof FIG. 3H indicates immobility times (sec) in the FST. (Example 2)

FIG. 3I is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the frontal cortex in the social defeat stressmice investigated 8 days after the injection of ketamine. In FIG. 3I,R-Ket and S-Ket represent social defeat stress mouse groups injectedwith R(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. In addition, mPFCmeans the medial prefrontal cortex. (Example 2)

FIG. 3J is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the hippocampal dentate gyrus in the socialdefeat stress mice investigated 8 days after the injection of ketamine.In FIG. 3J, R-Ket and S-Ket represent social defeat stress mouse groupsinjected with R(−)-ketamine and S(+)-ketamine, respectively. Salinerepresents a social defeat stress mouse group injected with saline, andControl represents a control mouse group injected with saline. (Example2)

FIG. 3K is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the hippocampus CAI region in the social defeatstress mice investigated 8 days after the injection of ketamine. In FIG.3J, R-Ket and S-Ket represent social defeat stress mouse groups injectedwith R(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. (Example 2)

FIG. 3L is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the hippocampus CA3 region in the social defeatstress mice investigated 8 days after the injection of ketamine. In FIG.3J, R-Ket and S-Ket represent social defeat stress mouse groups injectedwith R(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. (Example 2)

FIG. 3M is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the nucleus accumbens in the social defeatstress mice investigated 8 days after the injection of ketamine. In FIG.3J, R-Ket and S-Ket represent social defeat stress mouse groups injectedwith R(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. (Example 2)

FIG. 3N is a graph showing results of effects of R(−)- and S(+)-ketamineon the spine density of the striatum in the social defeat stress miceinvestigated 8 days after the injection of ketamine. In FIG. 3J, R-Ketand S-Ket represent social defeat stress mouse groups injected withR(−)-ketamine and S(+)-ketamine, respectively. Saline represents asocial defeat stress mouse group injected with saline, and Controlrepresents a control mouse group injected with saline. (Example 2)

FIG. 4 is a graph showing time-dependent changes in locomotion ofcontrol mice after the injection of R(−)- and S(+)-ketamine. In FIG. 4,R-Ket, S-Ket, and Saline represent groups injected with R(−)-ketamine,S(+)-ketamine, and saline, respectively. The ordinate axis of FIG. 4indicates locomotion (count/10min). (Example 3)

FIG. 5A is a graph showing changes in prepulse inhibition after theinjection of R(−)-ketamine in control mice. In FIG. 5A, R-Ket and Salinerepresent groups injected with R(−)-ketamine and saline, respectively.PP69, PP73, PP77, and PP81 mean that stimuli at 69, 73, 77, and 81 dBfor more than 20 milliseconds were presented 100 milliseconds before a110-dB pulse, respectively. Data analysis was performed by Wilks Lambda,which is multivariate analysis of variance. (Example 3)

FIG. 5B is a graph showing changes in prepulse inhibition after theinjection of S(+)-ketamine in the control mice. In FIG. 5B, S-Ket andSaline represent groups injected with S(+)-ketamine and saline,respectively. PP69, PP73, PP77, and PP81 mean that stimuli at 69, 73,77, and 81 dB for more than 20 milliseconds were presented 100milliseconds before a 110-dB pulse, respectively. Data analysis wasperformed by Wilks Lambda, which is multivariate analysis of variance.(Example 3)

FIG. 6A is a diagram illustrating a test protocol for investigatingrewarding effects of R(−)-ketamine, S(+)-ketamine, and RS(+/−)-ketamineon the control mice using a conditioned place preference test. 15-minutehabituation was performed for 3 days. Then, 30-minute conditioning wasperformed on day 4 to day 10, and a behavioral evaluation test wasperformed on day 11. Saline was injected on day 5, day 7, and day 9. InFIG. 6A, R-Ket, S-Ket, and RS-Ket mean groups injected withR(−)-ketamine, S(+)-ketamine, and RS(+/−)-ketamine, respectively. Ineach of the groups, the injection was performed three times, i.e., onday 4, day 6, and day 8. Saline means a group injected with saline. Inthe group, the injection was performed three times, i.e., on day 5, day7, and day 9. (Example 3)

FIG. 6B is a graph showing results of the rewarding effects ofR(−)-ketamine on the control mice using the conditioned place preferencetest. In FIG. 6B, R-Ketamine and Saline represent groups injected withR(−)-ketamine and saline, respectively. The ordinate axis of FIG. 6Bindicates conditioned place preference test scores (CPP scores).(Example 3)

FIG. 6C is a graph showing results of the rewarding effects ofS(+)-ketamine on the control mice using the conditioned place preferencetest. In FIG. 6C, S-Ketamine and Saline represent groups injected withS(+)-ketamine and saline, respectively. The ordinate axis of FIG. 6Cindicates conditioned place preference test scores (CPP scores).(Example 3)

FIG. 6D is a graph showing results of the rewarding effects ofRS(+/−)-ketamine on the control mice using the place preference test. InFIG. 6D, RS-Ketamine and Saline represent groups injected withRS(+/−)-ketamine and saline, respectively. The ordinate axis of FIG. 6Dindicates conditioned place preference test scores (CPP scores).(Example 3)

DESCRIPTION OF EMBODIMENTS

The present invention relates to an agent for prevention and/ortreatment of a depressive symptom, consisting of R(−)-ketamine or apharmacologically acceptable salt thereof. The present invention alsorelates to a pharmaceutical composition for prevention and/or treatmentof a depressive symptom, including R(−)-ketamine or a pharmacologicallyacceptable salt thereof in an effective amount for reducing a depressivesymptom, and being substantially free of S(+)-ketamine or apharmacologically acceptable salt thereof. The phrase “substantiallyfree of S(+)-ketamine or a pharmacologically acceptable salt thereof”means that: S(+)-ketamine or a pharmacologically acceptable salt thereofis not contained at all; or S(+)-ketamine or a pharmacologicallyacceptable salt thereof may be contained in such an amount that itseffects and side effects are not exhibited, or may be contained as suchan impurity as to be mixed inevitably during the manufacture of theagent and the pharmaceutical composition.

Further, the present invention relates to a method of treating adepressive symptom, comprising administering R(−)-ketamine or apharmacologically acceptable salt thereof to a subject, wherein thesubject has been diagnosed with having a depressive symptom, in anamount effective to treat the depressive symptom. The present inventionalso relates to a method of treating a depressive symptom, comprisingadministering a pharmaceutical composition to a subject, wherein thepharmaceutical composition comprises R(−)-ketamine or a pharmaceuticallyacceptable salt thereof in an amount effective to treat a depressivesymptom, and a pharmaceutically acceptable carrier, and beingsubstantially free of S(+)-ketamine or a pharmacologically acceptablesalt thereof, and wherein the subject has been diagnosed with having adepressive symptom.

Furthermore, the present invention also relates to use of R(−)-ketaminefor the manufacture of a pharmaceutical composition for treating adepressive symptom.

In the present invention, through the use of a new animal model ofdepression, it was demonstrated that R(−)-ketamine had rapid andlong-lasting antidepressant effects. The animal model was prepared bythe inventors of the present application based on their finding thatdepression-like behavior was found in mice exposed neonatally to DEX ata juvenile stage and an adult stage (Non Patent Literature 18; seeExample 1). The animal model exhibits depression-like behavior even at ajuvenile stage, and hence is useful as an animal model of depression inchildren as well as depression in adults.

In the present invention, it was also revealed that R(−)-ketamine hadantidepressant effects in a social defeat stress model as well. Thesocial defeat stress model is a typical animal model of depression,which is used throughout the world (Non Patent Literature 19).

R(−)-ketamine exhibited rapid and long-lasting antidepressant effects ondepression-like behaviors of the juvenile mice after the neonatal DEXexposure and the social defeat stress mouse model through its singleadministration (see Example 1 and Example 2). Meanwhile, inlocomotion-enhancing effects, disruption of prepulse inhibition, and adependence test using a conditioned place preference test, which areevaluation systems for side effects, significant changes were found inS(+)-ketamine, whereas such side effects were not found in R(−)-ketamine(see Examples 4, 5, and 6). In addition, in the conditioned placepreference test, RS(+/−)-ketamine increased a conditioned placepreference (CPP) score, indicating drug dependence of RS(+/−)-ketamine.Further, R(−)-ketamine has low affinity for the NMDA receptor ascompared to S(±)-ketamine, and thus is considered to have less sideeffects such as psychotomimetic effects. Accordingly, R(−)-ketamine canserve as a promising and safe antidepressant as compared toS(+)-ketamine and RS(+/−)-ketamine.

R(−)-ketamine or a pharmacologically acceptable salt thereof may be usedas an antidepressant, specifically, as an agent to be used for treatmentand/or prevention of depressive symptoms such as mood depression,lowering of motivation, anxiety, the accompanying insomnia and anorexia,and suicidal ideation.

The agent and pharmaceutical composition according to the presentinvention are preferably applicable to diseases exhibiting depressivesymptoms, for example, depression such as MDD or pediatric depression,and bipolar disorder involving a repeat of depressive symptoms and manicsymptoms as their opposite symptoms, and are more preferably applicableto depression in children and depression in adults. In addition, it hasbeen reported that ketamine is also effective for treatment-resistantobsessive-compulsive disorder and treatment-resistant PTSD (Non PatentLiteratures 6, 7, and 8). Thus, the agent and pharmaceutical compositionaccording to the present invention are preferably applicable toobsessive-compulsive disorder and PTSD. Obsessive-compulsive disorder,which is one type of anxiety disorder and is a disease with pathologicalconditions characterized by obsessions and compulsions, is considered tobe associated with depression. Patients with obsessive-compulsivedisorder have depression as well and exhibit depressive symptoms inaddition to obsessions and compulsions in extremely many cases. Patientswith PTSD exhibit depressive symptoms in many cases. In actuality, anantidepressant such as an SSRI is used as a therapeutic drug for PTSD,but its therapeutic effects are weak. The scope of the present inventionencompasses a pharmaceutical composition for prevention and/or treatmentof obsessive-compulsive disorder and PTSD, containing R(−)-ketamine or apharmacologically acceptable salt thereof in an effective amount forreducing symptoms of obsessive-compulsive disorder and PTSD, and beingsubstantially free of S(+)-ketamine or a pharmacologically acceptablesalt thereof. In addition, ketamine treatment in an adult with autismspectrum has been reported (Non Patent Literature 10). Thus, the agentand pharmaceutical composition according to the present invention arepreferably applicable to autism spectrum disorder. Furthermore,Depression in elderly peoples is known to increase the risk if incidentdementia, in particular of Alzheimer's disease and vascular dementia(Non Patent Literature 1). Therefore, the agent and pharmaceuticalcomposition according to the present invention is a potential preventiveor therapeutic drug for dementia including Alzheimer's disease andvascular dementia.

The agent and pharmaceutical composition according to the presentinvention may be administered orally or parenterally. In the oraladministration, a known dosage form for administration, including atablet, a capsule, a coated tablet, a troche, or a liquid such as asolution or a suspension, may be used. In addition, examples of theparenteral administration may include: intravenous, intramuscular, orsubcutaneous administration by injection; transmucosal administrationsuch as transnasal or oral administration using a spray, an aerosol, orthe like; rectal administration using a suppository or the like; andtransdermal administration using a patch, a liniment, a gel, or thelike. Preferred examples thereof may include oral administration,transnasal administration, and intravenous administration.

R(−)-ketamine may be used in both the forms of a free base and apharmaceutically acceptable salt thereof. The pharmaceuticallyacceptable salt is preferably a pharmaceutically acceptable acidaddition salt, more preferably a hydrochloride.

The chemical structural formula of R(−)-ketamine hydrochloride isrepresented by the following formula (I).

R(−)-ketamine or a pharmacologically acceptable salt thereof may besubjected to modification, for example, substitution of a chlorinemolecule as a substituent by another halogen molecule and/orsubstitution of a methyl group as a substituent by another alkyl group,to thereby manufacture a derivative. As a result, a compound having morepreferred effects may be obtained.

Further, when the compound according to the present invention is labeledwith an isotope such as a stable isotope ¹³C or ²H (D), the compound canbe measured for its in vivo kinetics and quantitatively measured for itsaffinity for the NMDA receptor in the brain, for example.

The pharmaceutical composition according to the present invention maycontain, in addition to R(−)-ketamine or a pharmacologically acceptablesalt thereof, other in having drug efficacy that are effective fordepressive symptoms, the ingredients gredients being other thanS(+)-ketamine. In addition, the pharmaceutical composition according tothe present invention may appropriately contain, in addition to thoseingredients having drug efficacy, an appropriate pharmaceuticallyacceptable carrier well known to those of ordinary skill in the art,depending on an administration form and the like. Examples of thepharmaceutically acceptable carrier may include an antioxidant, astabilizer, a preservative, a taste-masking agent, a colorant, asolubilizer, a solubilizing agent, a surfactant, an emulsifier, anantifoaming agent, a viscosity adjustor, a gelling agent, an absorptionaccelerator, a dispersant, an excipient, and a pH adjustor.

When the agent and pharmaceutical composition according to the presentinvention are each prepared as a formulation for injection, it ispreferred that the formulation be in the form of a solution or asuspension. When the agent and pharmaceutical composition are eachprepared as a formulation for transmucosal administration such astransnasal or oral administration, it is preferred that the formulationbe in the form of a powder, a drop, or an aerosol. In addition, when theagent and pharmaceutical composition are each prepared as a formulationfor rectal administration, it is preferred that the formulation be inthe form of a semi-solid formulation such as a cream or a suppository.Each of those formulations may be prepared by any one of the methodsknown to those skilled in the art of pharmacy as disclosed in, forexample, Remington's Pharmaceutical Sciences (Mack Publishing Company,Easton, Pa., 1970). In the formulation for injection, for example, aplasma-derived protein such as albumin, an amino acid such as glycine,and a sugar such as mannitol may each be added as a carrier, and abuffer, a solubilizing aid, an isotonic agent, and the like may also beadded. In addition, when the formulation is used as a water-solubleformulation or a lyophilized formulation, it is preferred to add asurfactant such as Tween™ 80 or Tween™ 20 in order to preventaggregation. Further, a dosage form for parenteral administration otherthan the formulation for injection may contain distilled water orsaline, polyalkylene glycol such as polyethylene glycol, a plant-derivedoil, hydrogenated naphthalene, and the like. For example, a formulationfor rectal administration such as a suppository contains generalexcipients such as polyalkylene glycol, petrolatum, and cacao oil andfat. A vaginal formulation may contain an absorption accelerator such asa bile salt, an ethylenediamine salt, and a citric acid salt. Aformulation for inhalation may be solid, and may contain an excipientsuch as lactose. Further, a transnasal drop may be a water or oilsolution.

The accurate dosage and dosing regimen of each of the agent andpharmaceutical composition according to the present invention may beadjusted depending on required amounts, treatment methods, diseases,degrees of necessity, or the like for individual treatment targets. Thedosage may be specifically deteimined depending on an age, a bodyweight, a general health condition, a sex, a meal, an administrationtime, an administration method, an elimination rate, a combination ofdrugs, a medical condition of a patient, and the like, and may bedetermined in consideration of other factors. When the pharmaceuticalcomposition according to the present invention is administered fordiseases exhibiting depressive symptoms, such as depression, bipolardisorder, and obsessive-compulsive disorder, it is preferred that anactive ingredient contained in the pharmaceutical composition becontained in an effective amount for reducing symptoms of the diseasessuch as depression, bipolar disorder, and obsessive-compulsive disorder,preferably depressive symptoms of the diseases. R(−)-ketamine or apharmaceutically acceptable salt thereof can be safely used because ofhaving less side effects found in S(+)-ketamine and RS(+/−)-ketamine.Its dosage per day varies depending on the condition and body weight ofa patient, the kind of a compound, an administration route, and thelike. For example, in terms of the amount of an active ingredient, it isdesired that the dosage in the case of parenteral administration be fromabout 0.01 to 1,000 mg/person/day, preferably from 0.1 to 500mg/person/day, and the dosage in the case of oral administration be fromabout 0.01 to 500 mg/person/day, preferably from 0.1 to 100mg/person/day.

EXAMPLES

The present invention is hereinafter described in more detail by way ofExamples. However, the present invention is by no means limited toExamples below. Further, various modifications are possible withoutdeparting from the technical concept of the present invention.

Example 1

A new animal model of depression (Non Patent Literature 18) was used toinvestigate antidepressant effects of R(−)- and S(+)-ketamine on thedepression-like behavior of the animal model. All tests were performedunder the approval of the Animal Care and Use Committee of ChibaUniversity.

1. Materials and Methods R(−)- and S(+)-ketamine hydrochloride wereprepared from RS(+/−)-ketamine (Ketalar™, ketamine hydrochloride.Daiichi Sankyo Co., Ltd., Tokyo, Japan) using D(−)- and L(+)-tartaricacid, respectively, by the method disclosed in the previous report(Patent Literature 2) (FIG. 1). The purity of each of those isomers wasconfirmed by high-performance liquid chromatography (CHIRALPAK™IA,column size: 250×4.6 mm, mobile phase;n-hexane/dichloromethane/diethylamine (75/25/0.1), retention time forS(+)-ketamine =6.99 min. retention time for R(−)-ketamine=10.56 min,Daicel Corporation, Tokyo, Japan).

A new animal model of depression was prepared by exposing miceneonatally to dexamethasone (hereinafter abbreviated as DEX). Throughthe neonatal DEX exposure, depression-like behavior was observed in eachof juvenile mice and adult mice. Thus, the mouse model was shown to beable to serve as a novel animal model of depression. The mouse model wasprepared and reported only recently by the inventors of the presentapplication and their collaborators (Non Patent Literature 18).Specifically, the juvenile mice exposed neonatally to DEX and the adultmice exposed neonatally to DEX showed a significant decrease in novelobject search time in a novel object recognition test as compared tocontrol mice, which indicated a reduction in social learning property inthe model mice. In addition, in a social memory test, the mice exposedneonatally to DEX showed a significant decrease in stimulation targetfollow-up time, which indicated a reduction in social recognitionability. In an open field test, a time spent in the center of a fieldsignificantly decreased, which indicated a reduction in spontaneousactivity. In a light-dark box test, a time spent in a white boxsignificantly decreased, which indicated that anxiety-like behavior wascaused. In each of a tail suspension test (TST) and a forced swimmingtest (FST), an increase in immobility time was found, which indicatedthat depression-like behavior was shown. Meanwhile, in a locomotion test(LMT), there was no difference in locomotion between the mice exposed toDEX and the control mice. Further, alterations were found in levels ofamino acids (glutamate, glutamine, glycine, D-serine, and L-serine) inmouse brains after the neonatal DEX exposure (Non Patent Literature 18).Those amino acids are known to be associated with NMDA receptor mediatedneurotransmission. Thus, it is conceivable that alterations inglutamatergic transmission via the NMDA receptor after the neonatal DEXexposure may be involved in the depression -like behavior in thejuvenile mice and the adult mice (Non Patent Literature 18).

The preparation of the animal model of depression and the administrationof the agent were specifically performed as described below (FIG. 2A).Male and female ICR mice (9-week-old, Japan SLC, Inc., Hamamatsu, Japan)were used. The mice were given free access to water and feed. A breedingprocedure consisted of housing three to four females with one male, for14 days. On the final day of this period, the females were placed inisolation and checked daily around the expected delivery day. The day ofbirth was defined as day 0. The mice were injected intraperitoneallywith DEX (Wako Pure Chemical Industries, Ltd., Tokyo, Japan) dissolvedin saline on day 1, day 2, and day 3 at doses of 0.5 mg/kg body weight,0.3 mg/kg body weight, and 0.1 mg/kg body weight, respectively. Inaddition, normal controls were injected with equal volumes (10 ml/kg) ofsaline. R(−)- or S(+)-ketamine at a dose of 10 mg/kg body weight orvehicle (saline 10 ml/kg) was injected intraperitoneally into malejuvenile mice on day 36 after the birth.

The antidepressant effects of the agent were investigated for juvenilemice by behavioral tests such as the TST, the FST, the LMT, and a 1%sucrose preference test (SPT) (FIG. 2A). The TST and the FST wereperformed twice, i.e., the day (27 hours and 29 hours, respectively) and7 days after the injection of ketamine, and the LMT and the SPT wereperformed on the day and 2 days after the injection of ketamine. The TSTwas perfoimed as described below. First, the mice were taken out fromcages, and then a small piece of an adhesive tape was bonded onto aportion approximately 2 cm away from the tip of the tail of the mice. Asmall hole was opened in the small piece, and the mice were each fixedupside down on a hook through the small hole. The immobility time ofeach mouse was recorded for 10 minutes. Mice were considered immobileonly when they hung passively and completely motionless. The immobilitytime increases in a depressive state. The FST was performed as describedbelow. First, the mice were placed individually in a cylinder (diameter:23 cm; height: 31 cm) containing 15 cm of water, maintained at 22 to 24deg C. The mice were tested in an automated forced-swimming apparatususing SCANET MV-40 (MELQUEST Co., Ltd., Toyama, Japan). The immobilitytime was calculated as a value obtained by subtracting active time fromtotal time, using the analysis software of the apparatus. Cumulativeimmobility time was recorded over 6 minutes during a test period. TheLMT was performed as described below. First, the mice were placed inexperimental cages (length×width×height: 560×560×330 mm). The locomotoractivity of the mice was counted with SCANET MV-40, and the cumulativeexercise of the mice was recorded for 60 minutes. The cages were cleanedbetween testing session. The immobility time increases in a depressivestate. The SPT was performed by preparing general drinking water and a1% sucrose solution so that the mice had free access thereto, andmeasuring the ratio of the amount of the sucrose solution consumed. Theconsumption of the sucrose solution, which is a reward response, reducesin a depressive state.

Statistical analysis was performed by one-way analysis of variance(one-way ANOVA), followed by a least significant difference test (LSDtest). Data are presented as the mean plus minus standard error of themean (n=8 to 12 mice/group). *p<0.05, ** p<0.01, and ***p<0.001 indicatesignificant differences as compared to a DEX-treated mouse groupinjected with saline, and #p<0.05 and ##p<0.01 indicate significantdifferences as compared to a DEX-treated mouse group injected withS(+)-ketamine.

2. Results

Significant increases in immobility time in the TST and the FST and areduction in sucrose consumption preference in the SPT were found in themice exposed neonatally to DEX as compared to the control mice. On theother hand, in the LMT, there was no difference in locomotion betweenthe DEX-treated mice and the control mice.

In the LMT performed on the day after the injection of both the isomersof ketamine, there was no difference in locomotion among the controlmice, the DEX-treated mice injected with saline, and the DEX-treatedmice injected with R(−)- or S(+)-ketamine (FIG. 2B).

In the TST and FST performed on the day after the injection of both theisomers of ketamine, significant increases in immobility time were foundin the DEX-treated mice injected with saline as compared to the controlmice. Each of both the isomers of ketamine markedly reduced theimmobility time increased in the DEX-treated mice 27 hours or 29 hoursafter its injection (FIGS. 2C and 2D). R(−)-ketamine exhibited slightlyhigh antidepressant effects as compared to those of S(+)-ketamine,although no significant difference was found there between.

In the SPT performed on 2 days after the injection of both the isomersof ketamine, a reduction in sucrose consumption preference was found inthe DEX-treated mice injected with saline as compared to the controlmice. Both the isomers of ketamine significantly restored the sucroseconsumption preference reduced in the DEX-treated mice 48 hours aftertheir injection (FIG. 2E).

In the TST and FST performed on 7 days after the injection of both theisomers of ketamine, significant increases in immobility time were foundin the DEX-treated mice injected with saline as compared to the controlmice. In addition, R(−)-ketamine significantly reduced the immobilitytime increased in the DEX-treated mice, whereas S(+)-ketamine did notreduce the immobility time increased in the DEX-treated mice. Thedifferences between R(−)-ketamine and S(+)-ketamine were found to bestatistically significant (FIGS. 2F and 2G).

The above-mentioned results revealed that R(−)- and S(+)-ketamine at adose of 10 mg/kg exhibited antidepressant effects in the juvenile miceafter the neonatal DEX exposure (days 1 to 3). In the TST and the FST,the antidepressant effects of both the isomers of ketamine were found 27to 29 hours after their single injection. It is noteworthy that in theTST and the FST, the antidepressant effects of R(−)-ketamine were ableto be detected even 7 days after its single injection, whereas theantidepressant effects of S(+)-ketamine were not able to be detected 7days after its single injection. The results show that R(−)-ketamine hasmore long-lasting antidepressant effects than S(+)-isomer. Both theisomers of ketamine are known to exhibit a rapid in vivo clearance.Despite the fact that R(−)-ketamine is considered to be eliminated fromthe body by 7 days after its single injection, the antidepressanteffects were found. This indicates that the differences inantidepressant effects 7 days after the injection of both the isomers ofketamine do not result from differences in pharmacokinetics.

Example 2

A social defeat stress model of depression (Non Patent Literature 19)was used to investigate antidepressant effects of R(−)- andS(+)-ketamine on the depression-like behavior of the animal model. Alltests were performed under the approval of the Animal Care and UseCommittee of Chiba University.

1. Materials and Methods

R(−)- and S(+)-ketamine hydrochloride were prepared fromRS(+/−)-ketamine (Ketalar™, ketamine hydrochloride, Daiichi Sankyo Co.,Ltd., Tokyo, Japan) using D(−)- and L(+)-tartaric acid, respectively, bythe method disclosed in the previous report (Patent Literature 2) (FIG.1). The purity of each of those isomers was confirmed byhigh-performance liquid chromatography (CHIRALPAK™ IA, column size:250×4.6 mm, mobile phase; n-hexane/dichloromethane/diethylamine(75/25/0.1), retention time for S(+)-ketamine=6.99 min, retention timefor R(−)-ketamine=10.56 min, Daicel Corporation, Tokyo, Japan).

A social defeat stress model of depression was prepared by bringingC57/B6 male mice into contact with ICR male mice (large aggressive mice)for 10 consecutive days to apply a stress called a “social defeatstress” in accordance with the previous report (Non Patent Literature19). Depression-like behavior was observed in the mice that had receivedthe social defeat stress. Specifically, an increase in immobility timewas found in each of a tail suspension test (TST) and a forced swimmingtest (FST). In addition, in a 1% sucrose preference test, the ratio ofsucrose water drunk significantly reduced, suggesting thatdepression-like behavior (e.g. anhedonia) was shown. On the other hand,in a locomotion test (LMT), there was no difference in locomotionbetween social defeat stress mice and control mice.

The preparation of the animal model of depression and the administrationof the agent were specifically performed as described below (FIG. 3A).Male C57/B6 mice (7-week-old, Japan SLC, Inc., Hamamatsu, Japan) and ICRmice (9-week-old, Japan SLC, Inc., Hamamatsu, Japan) were used. The micewere given free access to water and feed. A social defeat stress wasapplied by housing one C57/B6 mouse with one ICR mouse for 10 days. Onday 11, a social interaction test was performed to select miceexhibiting depressive symptoms, which were used for the subsequentbehavioral evaluation. Control mice were injected with vehicle (saline10 ml/kg) and the mice exhibiting depressive symptoms were injectedintraperitoneally with R(−)- or S(+)-ketamine at a dose of 10 mg/kg bodyweight, or vehicle (saline 10 ml/kg).

The antidepressant effects of the agent were investigated by behavioraltests such as the TST, the FST, the LMT, and a 1% sucrose preferencetest (SPT) (FIG. 3A). The 1% sucrose preference test (SPT) was performedon 1 day and 6 days after the injection of ketamine. Each of the TST andthe FST was performed on 2 days and 7 days after the injection ofketamine. The TST was performed as described below. First, the mice weretaken out from cages, and then a small piece of an adhesive tape wasbonded onto a portion approximately 2 cm away from the tip of the tailof the mice. A small hole was opened in the small piece, and the micewere each fixed upside down on a hook through the small hole. Theimmobility time of each mouse was recorded for 10 minutes. Mice wereconsidered immobile only when they hung passively and completelymotionless. The immobility time increases in a depressive state. The FSTwas performed as described below. First, the mice were placedindividually in a cylinder (diameter: 23 cm; height: 31 cm) containing15 cm of water, maintained at 22 to 24 deg C. The mice were tested in anautomated forced-swimming apparatus using SCANET MV-40(MELQUEST Co.,Ltd., Toyama, Japan). The immobility time was calculated as a valueobtained by subtracting active time from total time, using the analysissoftware of the apparatus. Cumulative immobility time was recorded over6 minutes during a test period. The LMT was performed as describedbelow. First, the mice were placed in experimental cages(length×width×height: 560×560×330 mm). The locomotor activity of themice was counted with SCANET MV-40, and the cumulative exercise of themice was recorded for 60 minutes. The cages were cleaned between testingsession. The immobility time increases in a depressive state. The SPTwas performed by preparing general drinking water and a 1% sucrosesolution so that the mice had free access thereto, and measuring theratio of the amount of the sucrose solution consumed. The consumption ofthe sucrose solution, which is a reward response, reduces in adepressive state. The mice were decapitated 5 days after the injectionof ketamine, and the brain was quickly dissected out and subjected toGolgi staining. A spine density was quantitatively evaluated byobservation with a KEYENCE microscope (BZ-9000, Osaka, Japan).

The statistical analysis of the results of the social defeat stressmodel was performed by one-way analysis of variance (one-way ANOVA),followed by a least significant difference test (LSD test). Data arepresented as the mean plus minus standard error of the mean (n=8 to 11mice/group). *p<0.05, **p<0.01, and ***p<0.001 indicate significantdifferences as compared to a social defeat stress mouse group injectedwith saline, and #p<0.05 indicates a significant difference as comparedto a social defeat stress mouse group injected with S(+)-ketamine.

2. Results

Significant increases in immobility time in the TST and the FST and asignificant reduction in sucrose consumption preference in the SPT werefound in the social defeat stress mice as compared to the control mice.On the other hand, in the LMT, there was no difference in locomotionbetween the social defeat stress mice and the control mice.

In the SPT performed on 1 day after the injection of both the isomers ofketamine, in the social defeat stress mice, the sucrose consumptionpreference significantly decreased as compared to the control group, anddepressive symptoms were exhibited. In the social defeat stress mousegroup injected with R(−)- or S(+)-ketamine, the sucrose consumptionpreference significantly increased as compared to the social defeatstress mouse group injected with saline, and depressive symptoms werealleviated. In addition, the antidepressant effects of R(−)-ketaminewere more potent than those of S(+)-ketamine (FIG. 3B).

In the LMT performed on 2 days after the injection of both the isomersof ketamine, there was no difference in locomotion among the normalmice, the social defeat stress mice injected with saline, and the socialdefeat stress mice injected with R(−)- or S(+)-ketamine (FIG. 3C).

In the TST and FST performed on 2 days after the injection of both theisomers of ketamine, significant increases in immobility time were foundin the social defeat stress mice injected with saline as compared to thecontrol mice. Each of both the isomers of ketamine markedly reduced theimmobility time increased in the social defeat stress mice 2 days afterits injection (FIGS. 3D and 3E). R(−)-ketamine exhibited significanthigh antidepressant effects as compared to those of S(+)-ketamine (FIGS.3D and 3E).

In the SPT performed on 6 days after the injection of both the isomersof ketamine, a reduction in sucrose consumption preference was found inthe social defeat stress mice injected with saline as compared to thecontrol mice. Both the isomers of ketamine significantly restored thesucrose consumption preference reduced in the social defeat stress mice6 days after their injection. The difference between R(−)-ketamine andS(+)-ketamine was found to be statistically significant (FIG. 3F).

In the TST and FST performed on 7 days after the injection of both theisomers of ketamine, significant increases in immobility time were foundin the social defeat stress mice injected with saline as compared to thecontrol mice. Each of both the isomers of ketamine significantly reducedthe immobility time increased in the social defeat stress mice 7 daysafter its injection (FIGS. 3G and 3H). R(−)-ketamine exhibitedsignificantly high antidepressant effects as compared to S(+)-ketamine(FIGS. 3G and 3H).

In the Golgi staining performed 8 days after the injection of both theisomers of ketamine, a significant decrease in spine density was foundin the frontal cortex and hippocampal dentate gyms of the social defeatstress mice injected with saline as compared to the control mice. Eachof both the isomers of ketamine significantly improved the decreaseddensity of spine in the social defeat stress mice 8 days after itsinjection (FIGS. 3I and 3J). In the hippocampal dentate gyrus,R(−)-ketamine showed more significant improvement in the restoration ofspine density as compared to SW-ketamine (FIG. 3J). In the hippocampusCAI region and striatum, no apparent change in spine density wasobserved (FIGS. 3K and 3N), while in hippocampus CA3 region, a decreasein spine density by social defeat stress was observed, which wassignificantly improved by R(−)-ketamine and SW-ketamine (FIG. 3L). Inthe nucleus accumbens, a significant increase in spine density wasobserved by social defeat stress, which was significantly improved byR(−)-ketamine and S(+)-ketamine (FIG. 3M).

The above-mentioned results revealed that R(−)- and SW-ketamine at adose of 10 mg/kg exhibited antidepressant effects in the social defeatstress mice. It is noteworthy that in the SPT, the TST, and the FST, theantidepressant effects of R(−)-ketamine were significantly potent ascompared to the effects of SW-ketamine. The results show thatR(−)-ketamine has more long-lasting antidepressant effects thanSW-isomer. Both the isomers of ketamine are known to exhibit a rapid invivo clearance. Despite the fact that R(−)-ketamine is considered to beeliminated from the body by 7 days after its single injection, theantidepressant effects were found. This indicates that the differencesin antidepressant effects 7 days after the injection of both the isomersof ketamine do not result from differences in pharmacokinetics.

Example 3

Control C57/B6 mice was used to investigate side effects of R(−)- andSW-ketamine. All tests were performed under the approval of the AnimalCare and Use Committee of Chiba University.

1. Materials and Methods

The preparation of R(−)- and SW-ketamine hydrochloride and theconfirmation of their purities were performed by the methods describedin Example 2.

The administration of an agent was performed by the same methods as themethods described in Example 2.

The side effects of R(−)- and S(+)-ketamine were investigated by alocomotion-enhancing effect, disruption of prepulse inhibition, and adependence test using a conditioned place preference test, which weresystems for evaluating side effects.

An effect of ketamine on the locomotion of mice was tested using SCANETMV-40 (MELQUEST Co., Ltd., Toyama, Japan). Specifically, the locomotionwas measured for a total of 180 minutes, i.e., 60 minutes beforeinjection to 120 minutes after injection, and calculated as a locomotionper 10 minutes. The statistical analysis of the results of thelocomotion was performed by repeated one-way analysis of variance(repeated one-way ANOVA), followed by a least significant differencetest (LSD test). Data are presented as the mean plus minus standarderror of the mean (n=7 or 8 mice/group). **p<0.01 and ***p<0.001indicate significant differences as compared to a group injected withsaline.

The prepulse inhibition test was performed using a startle responsesystem (SR-LAB, SanDiego Instruments, San Diego, Calif., United States).The analysis of the results of prepulse inhibition was performed bymultivariate analysis of variance (MANOVA), followed by a leastsignificant difference test (LSD test). Data are presented as the meanplus minus standard error of the mean (n=10 to 12 mice/group). *p<0.05and ** p<0.01 indicate significant differences as compared to a groupinjected with saline.

The place preference test was performed using a conditioned placepreference test apparatus (BrainSienceldea Co., Ltd., Osaka, Japan). Theanalysis of the results of the place preference test was performed byone-way analysis of variance (one-way ANOVA), followed by a leastsignificant difference test (LSD test). Data are presented as the meanplus minus standard error of the mean (n=9 or 10 mice/group). *p<0.05and **p<0.01 indicate significant differences as compared to a groupinjected with saline.

2. Results

In the measurement of the locomotion after the injection of both theisomers of ketamine, a significant increase in locomotion was found 10minutes and 20 minutes after the injection in the mice injected withS(+)-ketamine (10 mg/kg or 20 mg/kg) as compared to the control miceinjected with saline. The locomotion was transiently enhanced byS(+)-ketamine (10 mg/kg or 20 mg/kg). but returned to a normal value 30minutes after the injection. On the other hand, the injection ofR(−)-ketamine (5, 10, or 20 mg/kg) did not affect the locomotion (FIG.4).

In the prepulse inhibition test after the injection of both the isomersof ketamine, the injection of S(+)-ketamine (5, 10 mg/kg, or 20 mg/kg)disrupted prepulse inhibition in a dose-dependent manner (FIG. 5B). Onthe other hand, the injection of R(−)-ketamine (5, 10, or 20 mg/kg) didnot disrupt prepulse inhibition (FIG. 5A).

In the conditioned place preference test after the injection of both theisomers and racemic mixture of ketamine, the injection of S(+)-ketamine(5, 10 mg/kg, or 20 mg/kg) increased the CPP score in a dose-dependentmanner, indicating drug abuse potential (FIG. 6C). On the other hand,the injection of R(−)-ketamine (5, 10, or 20 mg/kg) did not increase theCPP score (FIG. 6B), indicating no drug abuse potential. In addition,the injection of RS(+/−)-ketamine (10 mg/kg) significantly increased theCPP score, indicating drug abuse potential (FIG. 6D).

As described above, from the viewpoint of side effects, the injection ofS(+)-ketamine was found to exhibit a locomotion-enhancing effect,disrupt prepulse inhibition, and produce drug dependence. In addition,it was suggested that the injection of RS(+/−)-ketamine also produceddrug dependence. On the other hand, R(−)-ketamine does not exhibit alocomotion-enhancing effect, disrupt prepulse inhibition, and producedrug dependence, for example, and hence is an agent having high safetyas compared to RS(+/−)-ketamine and S(+)-ketamine that are clinicallyused at present.

INDUSTRIAL APPLICABILITY

As described above, the agent and pharmaceutical composition forprevention and/or treatment of a depressive symptom according to thepresent invention have rapid and long-lasting antidepressant effects andless side effects such as psychotomimetic effects, and hence are usefulas novel pharmaceuticals in the field of prevention and/or treatment ofa number of psychiatric diseases exhibiting depressive symptoms.

1-18. (canceled)
 19. A method of treating a depressive symptom,comprising administering a derivative of R(−)-ketamine or apharmacologically acceptable salt thereof to a subject, wherein thederivative of R(−)-ketamine is represented by Formula (I):

wherein the chlorine molecule is substituted for another halogenmolecule, the methyl group is substituted for another alkyl group, or acombination thereof.
 20. The method of claim 19, wherein the derivativeof R(−)-ketamine is substantially free of S(+)-ketamine, a derivative ofS(+)-ketamine, or a pharmacologically acceptable salt thereof.
 21. Themethod of claim 20, wherein the derivative of R(−)-ketamine comprises noS(+)-ketamine, or the pharmacologically acceptable salt thereof.
 22. Themethod of claim 20, wherein the derivative of R(−)-ketamine comprises anamount of S(+)-ketamine, the derivative or pharmacologically acceptablesalt thereof such that side effects of the S(+)-ketamine, derivative orthe pharmacologically acceptable salt thereof are not exhibited.
 23. Themethod of claim 22, wherein the side effects comprise psychotomimeticeffects.
 24. The method of claim 19, wherein the subject has beendiagnosed with having a depressive symptom.
 25. The method of claim 19,wherein the subject has depression, obsessive-compulsive disorder,posttraumatic stress disorder, major depressive disorder, bipolardisorder, dementia, suicidal ideation, low motivation, mood depression,anxiety, insomnia, or anorexia.
 26. The method of claim 25, wherein thedementia is Alzheimer's disease, vascular dementia, or a combinationthereof.
 27. The method of claim 19, wherein the subject is a child oran adult.
 28. The method of claim 27, wherein the adult is an elderlyadult.
 29. The method of claim 19, wherein the derivative ofR(−)-ketamine is administered in a therapeutically effective amount totreat the depressive symptom.
 30. The method of claim 29, wherein thetherapeutically effective amount is about 0.01 mg/day to about 500mg/day, about 0.1 mg/day to about 500 mg/day, or about 0.1 mg/day toabout 100 mg/day.
 31. The method of claim 29, wherein the derivative ofR(−)-ketamine is therapeutically effective for 24 hours, 48 hours, 6days or 7 days after administration to the subj ect.
 32. The method ofclaim 19, wherein the derivative of R(−)-ketamine is formulated in apharmaceutical composition further comprising a pharmaceuticallyacceptable carrier.
 33. The method of claim 32, wherein thepharmaceutical composition is suitable for intravenous, intramuscular,subcutaneous, transnasal, oral, rectal or transdermal administration.34. The method of claim 34, wherein the derivative of R(−)-ketamine isformulated a liquid, solution, suspension, powder, tablet, coatedtablet, capsule, troche, cream, suppository, gel, patch, liniment oraerosol.
 35. A pharmaceutical composition, comprising a derivative ofR(−)-ketamine or a pharmacologically acceptable salt thereof, whereinthe derivative of R(−)-ketamine is represented by Formula (I):

wherein the chlorine molecule is substituted for another halogenmolecule, the methyl group is substituted for another alkyl group, or acombination thereof.
 36. The pharmaceutical composition of claim 35,wherein the derivative of R(−)-ketamine is substantially free ofS(+)-ketamine, a derivative of S(+)-ketamine, or a pharmacologicallyacceptable salt thereof.
 37. The pharmaceutical composition of claim 35,wherein the derivative of R(−)-ketamine comprises no S(+)-ketamine, orthe pharmacologically acceptable salt thereof.
 38. The pharmaceuticalcomposition of claim 35, wherein the derivative of R(−)-ketaminecomprises an amount of S(+)-ketamine, the derivative orpharmacologically acceptable salt thereof such that side effects of theS(+)-ketamine, derivative or the pharmacologically acceptable saltthereof are not exhibited.
 39. The pharmaceutical composition of claim35, wherein the pharmaceutical composition is suitable for intravenous,intramuscular, subcutaneous, transnasal, oral, rectal or transdermaladministration.
 40. The pharmaceutical composition of claim 35, whereinthe derivative of R(−)-ketamine is formulated a liquid, solution,suspension, powder, tablet, coated tablet, capsule, troche, cream,suppository, gel, patch, liniment or aerosol.
 41. A method of treating adepressive symptom, comprising administering R(−)-ketamine, a derivativeof R(−)-ketamine, or a pharmacologically acceptable salt thereof to asubject, wherein the subject has been diagnosed with having a depressivesymptom, in an amount effective to treat the depressive symptom, andbeing substantially free of S(+)-ketamine and pharmacologicallyacceptable salts thereof, wherein the derivative of R(−)-ketamine isrepresented by Formula (I):

wherein the chlorine molecule is substituted for another halogenmolecule, the methyl group is substituted for another alkyl group, or acombination thereof.
 42. The method of claim 41, wherein the chlorinemolecule of Formula (I) is substituted for another halogen molecule. 43.The method of claim 41, wherein the methyl group of Formula (I) issubstituted for another alkyl group.
 44. The method of claim 41, whereinthe chlorine molecule of Formula (I) is substituted for another halogenmolecule, and the methyl group of Formula (I) is substituted for anotheralkyl group.